The molecular markers and bioinformatics were used to identify the genotypes and combinations of the F-M locus, transposon insertion in YUC11 promoter region, and 9 bp insert in NAC72 coding region by 189 peach germplasms, in order to provide the theoretical foundation for the mechanism of peach flesh formation and the selection of breeding parents.

PCR amplification, KASP, and HRM were used to identify genotypes. Results of loci associated with the formation of flesh trait were further validated using bioinformatics by resequencing data in 189 peach germplasms.

Two genes encoding endopolygalacturonase in F-M locus, designated PGM and PGF, are associated with peach texture. Through primer amplification, 159 (84%) PGM and 99 (52%) PGF were detected in 189 peach germplasms. Four haplotypes (H0, H1, H2, and H3) were found in F-M locus, while H0 and H1 were the major genotypes. The haplotype combination of non-melting flesh, melting flesh, free-stone melting flesh were H3H3 and H2H3, H0H0 and H0H1, H1H1 and H0H1, respectively. In addition, it was found that 18 germplasms contain homozygous transposon insertion. HRM results demonstrated that 45 germplasms were homozygous insertion (early ripening), 71 germplasms were heterozygous insertion (middle ripening), and 73 germplasms were no insertion (late ripening). But six germplasm genotypes were inconsistent with the phenotypes. The most frequent genotypic combinations at the three loci of non-melting flesh, melting flesh, and free-stone melting flesh were mmffHdHdI, MMffHdHdI, MMFFHdHdL, respectively.

The existence of four haplotypes at the F-M locus further was confirmed, 18 germplasms containing stony hard genotype were identified, a molecular marker for the identification of maturity date was developed, and genotype combinations of different germplasms at flesh-related loci were identified.